Design, construction, and application of a device for obtaining radiographs of microscopic objects in a commercial model electron microscope



July 15, 1958 M. c. BOTTY ET AL DESIGN, CONSTRUCTION AND APPLICATION OF A DEVICE FOR OBTAINING RADIOGRAPHS OF MICROSCOPIC OBJECTS IN A COMMERCIAL MODEL ELECTRON MICROSCOPE 2 Sheets-Sheet 1 Filed Feb. 6, 1956 ELECTRON F/LAMENT y A/VODE; i

FOCAL PO/N 0 OF L ENS \SPECIMEN IN CONTACT WITH FILM l l I l I I CONDENSER L EN$ TARGET AT FOCAL POINT OF LENS POL E P/E CE INVENTORS. MART/IV C. BOTTY BY FREDERICK 6. R0

ATTORNEY.

2 Sheets-Sheet 2 M. c. BOTTY ET AL DESIGN. CONSTRUCTION AND APPLICATION OF A DEVICE FOR OBTAINING RADIOGRAPHS OF MICROSCOPIC OBJECTS IN A COMMERCIAL MODEL ELECTRON MICROSCOPE Jul 15, 1958 Filed Feb. 6, 1956 INVENTORS.

MART/N 6. 8077') FREDERICK G. ROWE ATTORNEY DESIGN, CONSTRUCTION, AND APPLICATION OF A DEVICE FOR OBTAHNHNG RADTOGRAPHS OF MICROSCOPIC OBJECTS TN A COB/EMERQEAL MODEL ELECTRON MECROSOTE Martin Charles Betty, Norwallr, and Frederick George Rowe, Darren, Conan, assignors to American Cyanamid Company, New York, N. ifi, a corporation of Maine Application February d, 1956, Serial No. 563,466

4 Claims. Cl. 250*65) This invention relates to a combined electron and X-ray microscope and to an adapter usable with standard electron microscopes for transforming them into X-ray microscopes.

Microscopy has been faced with two types of problems, namely, resolutionand specimen opacity. The ordinary light microscope is limited in its resolution by the relatively long wave length of the radiant energy used and even the use of ultraviolet light extends but little the degree of resolution possible. The problem of resolution was brilliantly solved by the electron microscope in which a stream of electrons takes the place of the radiant energy. 'It behaves as if it had an effective or equivalent wave length far shorter than any visible or ultraviolet radiation and is limited only by the practical accelerating voltages which can be applied to the electron beam. Resolution orders of magnitude greater than obtainable with visible or ultraviolet light microscopes are readily obtained.

The second problem, specimen opacity, is not too serious in ordinary microscopy where the light is transmitted differentially through the specimen or where, as in electron microscopy, there is obtained an image which might be considered very largely as a silhouette. in electron microscopy, which is practically-exclusively a transmission phenomenon, and in certain phases of ordinary microscopy, which will be discussed below, opacity of the specimen becomes a controlling factor which limits the use of the technique. Most substances absorb electrons so easily that specimens must be extremely thin. The effective penetration of electronsin most solids and many liquids for useful resolution is of the order of not more than about 100 Angstrom units. This requires a very thin specimen and eliminates from .the field of electron microscopy 2. very large number of possible uses. in the case of ordinary lightmicrosc'opy, speciment opacity also comes into consideration with highly opaque particles. These par ticles can be seen only as silhouettes; that is to say, they contrast sharply from their background, but it is impossible to determine the interior structure of the particles because their surface layers are sufiiciently opaque so that in many cases of reflected light microscopy, one sees only the surface or, in the case of transmission samples, the boundary between particles of one material and other material surrounding it.

The bottleneck presented by specimen opacity could easily be solved if it were possible to use in a microscope more penetrating radiation such as X-rays of varying degree of hardness, i. e., of shorter or longer wave length.

, The use of X-rays, however, has hitherto been impractical for microscopy because of the difficulty of focusing or directing beams of X-rays. If it were possible to approach lengths.

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2 a point source of X-rays, proper positioning of a specimen would permit an enlargement or a contact image. The close approach to a point source produces beam conditions which are similar or analogous to those in a socalled pin-hole camera.

It was proposed to obtain an approximation of a point source by utilizing an electron microscope and focusing its electron beam as a fine point on a suitable metal target. Because the specimen must be illuminated by X-rays from the opposite side of the target, the latter must be an extremely thin metal foil as it must act both as a target for transformation of the beam of electrons into X-radiation and a window therefor. Serious structural problems are presented which will be discussed below in connection with one of the modifications of the present invention, and the nature of the target metal itself also imposes some other critical limitations. When most metals are bombarded with high-ener y electrons, two types of X-radiation result. One is peculiar to the metal itself and is in the form of one or more lines or sharp peaks in the X- ray spectrum analogous to emission or absorption lines of elements in the visual and ultraviolet. The other form of radiation is spread over a band of wave lengths without sharp peaks and is sometimes referred to as white radiation.

The characteristics of the radiation are effected both by the metal of the target and by the accelerating voltage which determines the velocity of the electrons striking the target. in general, lighter metals, such as aluminum, require a lower voltage to excite their characteristic radiation, and the wave length of their characteristic radiation is relatively long. Heavier metals, such as platinum, require a much higher voltage for excitation of characteristic radiation, and produce strong lines at much shorter wave The average energy distribution of the white radiation is also shifted to shorter wave lengths with higher voltage. In many cases the X-rays produced in a standard electron microscope with a high accelerating voltage of the order of 50 kv. are too hard, i. e., too short in wave length to give good contrast with many types of specimens.

It would be ideal if it were possible to use X-radiation in the form of the strongest characteristic line of a metal; but often, as pointed out above, this radiation is too hard, and, therefore, if low enough voltage is used, only the white radiation is obtained or this may constitute the major portion of the radiant energy. It is possible to produce the ideal line source by using a laminated target in which the lamination struck by the electrons produces X-rays which then excite the second lamination, which is normally a different metal to produce secondary X-rays by a phenomenon analogous to fluorescence; that is to say, where the second lamination transforms the shorter wave length radiation received into a line or narrow band of longer radiation. This type of laminated target is described in the Journal of Applied Physics, vol. 23, p. 881 et seq. Laminated targets, however, have the disadvantage that the intensity of the secondary X-rays is very much lower than the primary X-radiaticn and sometimes the efficiency is too low to permit practical operation. An

example of a lamina-ted fluorescent target would be copper, which gives off a characteristic radiation of wave length 1.54 A. under suitable excitation with a second lamination of chromium, which produces a characteristic radiation of about 2.23 A.

Another problem presented in the case of many specimens is the effect on the specimen of the environment during the microscopical examination. One of the factors is the effect of the energy on the specimen, electrons, in the case of the electron microscope, and X-rays, in the case of X-ray microscopy. Both result in a lethal effect on living micro-organisms. In addition, the electron stream may have sufficient energy to produce gross changes in fragile organisms, particularly in the extraordinarily thin specimens necessitated by the very low penetration of electrons. In many cases a more serious environmental problem is presented by the fact that, in electron microscopy and hitherto in X-ray work, the specimen was in the very high vacuum necessitated by the operating conditions of an electron microscope. This extraordinarily high vacuum resulted in far-going and very rapid desiccation of the specimen which frequently resulted in such physical changes as to destroy much of the value of the photomicrographs obtained, for they did not represent photographs of the specimen as it originally existed but only as it originally existed after being almost completely desiccated.

Another problem presented by radiographs produced in an electron microscope is the fact that the photographic emulsion is sensitive not only to electromagnetic radiation, such as the various X-rays, but also to moving charged particles, such as electrons, ions and the like. In some cases this presents a problem of false or background illumination of the photographic emulsion by moving charged particles in addition to the X-radiation, which, after passing through the specimen, is the only radiation carrying to the photographic film the desired information of specimen structure.

The present invention solves the practical problems peculiar to X-ray micrography and, of course, also to X- ray microscopy where a fluorescent screen takes the place of the photographic emulsion and includes a simple, economical and accurate device, which can be used with a standard electron microscope. Essentially the present invention provides a chamber in the pole piece, preferably of the second or objective lens, of the electron microscope. This device includes a thin X-ray target, a specimen holder, and a holder for photographic film or, if desired, a fluorescent screen. The chamber provides for an accurate positioning of the target at the focal point of the electron beam for accurate positioning of the specimen and of the film carrier. In the preferred modification of the present invention, this chamber is fabricated of non-magnetic, and preferably electrically conductive, material so that the focusing of the electron beam is not interfered with. It is removable and can be easily inserted into the standard type of pole piece used in electron microscopes. The chamber carries at its top the thin target foil, which can be as thin as a micron or less, preferably clamped between supporting surfaces so that only the portion of the target foil which is the actual X-ray source is left unsupported. Underneath is an accurately positionable specimen holder and a film holder, in the case of a chamber which is to be used for either contact or projection micrography.

In a more specific aspect of the present invention, the chamber is closed and can be filled with air or any other gas at a suitable pressure and with any desired degree of humidity and other air characteristics. In this more specific modification, specimens which are damaged by desiccation may be examined without destruction of their original form.

The invention will be described in greater detail in conjunction with the drawings in which:

Fig. l is a diagrammatic view of an electron microscope showing the beam of electron and X-radiation for projection radiography;

Fig. 2 is a similar diagram for contact radiography;

Fig. 3 is a vertical section through a pole piece;

Fig. 4 is a vertical section through one type of adapter, and

Fig. 5 is a vertical section through a closed-chamber adapter.

Fig. 1 illustrates, in diagrammatic form, the combination of a closed-chamber adapter and an electron microscope. Only the necessary elements defining the beams are shown, the other portions of the conventional electron microscope being omitted as they are not necessarily changed by the present invention. An electron-emitting filament or cathode 1 emits a stream of electrons which are accelerated by the anode 2 and condensed by the lens 3 and asecond lens, of which only the pole piece 4 is shown. Inside the pole piece at the focal point of the electron beam is positioned a thin, foil X-ray target 6 carried at the top of the chamber or adapter 5. Below the foil is positioned the specimen 7 and a film is shown at 8. The electron beam is shown in short dashes and the X-ray beam in dash and dot.

Fig. 2 illustrates the same set-up With the specimen in contact with the film. The same parts bear the same reference numerals. In the case of both Fig. 1 and Fig. 2 the representation is purely diagrammatic to show the travel of the beams as the elements of the electron microscope are conventional and the structural design of the adapter or chamber is shown and will be described in connection with Figs. 3 and 4. In Fig. 1 the specimen is shown separated a considerable distance from the target so that the X-ray beams can be shown clearly. This distance is exaggerated for the sake of illustration of the beams and actually should be very close to the target, generally a matter of a millimeter or even less.

It will be noted that in the combination illustrated in Figs. 1 and 2, Walls of the adapter constitute a chamber which shields the X-ray portion of the device from the electron microscope itself and which may be, as diagrammatically illustrated, completely closed so that the environment within the chamber or adapter can, if desired, be entirely different from the high vacuum existing in the pole piece of the electron microscope.

The operation of the combination of the present invention is simple. The electron microscope is adjusted to produce an electron beam of the desired velocity for the particular wave length range of X-radiation required. In other words, preferably the electron microscope is operated at different accelerating voltages, although it is possible to use the features of the present invention with an electron microscope having a fixed accelerating voltage. Of course, if the voltage is varied, the microscope must be adjusted so that the beam is focused at the proper point. With standard electron microscopes which often have a high accelerating voltage, for example of the order of 50 kv., the X-radiation produced is comparatively hard. For many specimens this is entirely satisfactory, but in the case of specimens of low contrast, that is to say where the different elements in the specimen do not have very high differences in density, little or no structural detail will be seen in the highlights. For example, some particles of metallized dyestulf of low density in a section of a fiber may show little contrast from the fiber itself. On the other hand, with specimens of greater thickness or containing elements of very high density such as, for example, specimens of ores, metals and the like, the hard X-rays give optimum results. Softer X-rays can be produced in electron microscopes which are provided with variable accelerating voltages or to which a variable power source is added. They permit examination of the widest range of specimens and for many purposes are preferable, although, of course, where all specimens to be examined require hard X-rays, an electron microscope with a single, high-accelerating voltage will give satisfactory results.

Fig. 3 illustrates a conventional pole piece Which has an upper part 29, a brass spacer 30 separating the lower portion 31 from the upper part. For simplicity the brass spacer is shown without centering screws for centering its aperture as these refinements of structure are not in any way changed by the use of the present invention.

Fig. 4 illustrates a simple chamber or adapter which is normally, although not necessarily, used in a vacuum insidethe pole piece. The adapter is provided with a positioning ring 9 into which the main body 10 of the chamber is screwed, which main body also carries a flange or enlarged portion 11 having approximately the outer diameter of the adjusting ring 9. This outer diameter is such that it provides a snug sliding fit in the inside of the lower portion 31 of the pole piece. The main body 10 is hollow and threaded on its inside to take a threaded sleeve 12 having a central opening 13 which contains the sliding specimen holder 14. The specimen is placed in the desired position below the foil target 6 which is at the upper end of the threaded sleeve 12. The film is applied in the enlarged portion 11 of the main body 10 and is therefore a fixed distance from the specimen, thus providing for a magnification. If it is desired to effect contact radiography, the specimen may be placed directly on the film at 11. If shorter exposure time is desired, the film may be placed within the main body 10 nearer to the target 6. The assembled device is inserted in the pole piece, Fig. 3, so that the target 6 coincides with the point source of electrons existing in the space in the vicinity of components 30 and 31 of Fig. 3.

Fig. 5 illustrates a more complex and flexible chamber or adapter. Here a main shell 15 is screwed into a bottom piece 16, and on it is screwed an upper conical portion 17, gaskets 32 assuring a gas-tight joint in each case. The upper portion 17 carries a top piece 18 which is threaded and which has a very small aperture in the center. This top carries shoulders 19 which clamp the target foil 21 against shoulders on the conical part 17, gaskets 32 being provided for gas-tightness, as in the other seals. The upper end of the chamber 15 also is internally threaded, and a hollow sleeve 22 screws into said thread. This sleeve has a depression 23 for holding the specimen.

In the lower part of the chamber 15 is mounted a film carrier 26 with a depression 24 for the film and a suitable slot 25 to facilitate removing the film after use. A lower portion of the same carrier communicates with the chamber 15 through the channels 27 and to the outside through a threaded opening in which the screw 28 can be inserted.

The adapter illustrated in Fig. 5 permits positioning of the specimen with respect to the target as in the device in Fig. 4 but it also permits maintaining a pre-determined environment, such as air or other gas, under pre-determined pressure within the chamber, formed by the e1ements 15 and 17. Therefore, the specimen is not exposed to the high vacuum in the pole piece when the electron microscope is being operated. At the same time if it is desired that a vacuum be present in the adapter containing the specimen, this is easily effected by removing the screw 28. It will be noted that the target 21 is clamped tightly between the shoulders 19 and 20 so that only the very tiny aperture is exposed to the difference in pressures existing between the electron microscope and the adapter chamber. Therefore, even when the chamber is at atmospheric pressure, or near to it, it is possible to use very thin foils as the target because the opening is of such small diameter that rupture of the foil by bending, which is a serious factor with larger foils, is practically eliminated. This modification therefore permits operating the device with a predetermined environment for the specimen which does not have to be in the high vacuum of the electron microscope, as was hitherto necessary.

A very accurate spacing of the target from the aperture of the pole piece 30 is provided by the length of the elements 17 and 18. Normally, it is sufficient to use a standard length suitable for average operating conditions and to move the focal point of the electron beam onto the target by suitable adjustments of the microscope lens. Once these adjustment have been made, the alignment is maintained because every time the adapter is inserted with a new specimen, the element 18 acts as an accurate spacer, locating the target at a predetermined place. As .in Figs. 1 and 2, the specimen-carrying depression 23 is shown at considerable distance from the target in order that the elements of the adapter can be shown more clearly. Actually, ofcourse, the spacing of the specimen from the target is much closer.

Fig. 3 shows a conventional type of pole piece for electron microscopes having electromagnetic lenses, and the shape of the adapter shown in Figs. 4 and 5 corresponds approximately to the inside dimensions of the pole piece in which they are to be used. Where the shape of the pole piece is different, the outer dimensions and shapes of the adapters must correspond because, unless there is a snug fit, the accurate alignment of target, specimen and film is not maintained.

The adapter shown in Fig. 5 has a further advantage in that it is a very easy matter to change target foils and that the target is reliably clamped in proper position. This ready interchangeability of foils is sometimes an advantage when changing from specimens which can endure the high vacuum to other specimens in which the modification of the present invention, permitting a different environment, is necessary. It is sometimes desirable to change target foils as, of course, an even thinner foil may be used where there is no pressure differential between its two sides. It is, however, an advantage of the present invention that the clamping and support of the adapter of the preferred modification permits operation with foils that are thin enough for most X-ray micrography, even though atmospheric pressure prevails in the chamber. In other words, for most uses, it is not necessary to change foils, which, of course, makes for an additional operating advantage. At the same time the use of different foils is made possible and repairs in the case of damage to a foil are readily performed.

We claim:

1. An adapter for transforming an electron microscope into an X-ray microscope comprising, in combination, a chamber adapted to be slidably introduced into the pole piece of the second lens of the microscope and comprising, in combination, a hollow chamber member, a second hollow member adjustably positioned therein along the longitudinal axis thereof, said inner member being provided with specimen-holding means and filmholding means separated by a pre-deternrined distance from each other.

2. An adapter for transforming an electron miscroscope into an X-ray microscope comprising, in combination, an elongated chamber dimensioned for a sliding fit in an electron microscope pole piece, said chamber provided at its upper end with clamping means for an X-ray target foil forming a minute disc of the target foil aligned with the longitudinal axis of the pole piece, specimen carrying means longitudinally adjustable in said chamber, film-supporting means carried by said chamber and means for retaining a pre-determined atmosphere within said chamber and in contact with said specimen.

3. A device according to claim 2 in which the chamber is in a plurality of sections comprising a lower section adapted to fit snugly in a pole piece of an electron microscope, an upper tapered portion carrying the foil-clamping means, an internally threaded shoulder within the lower portion of the chamber, hollow specimen carrying means threaded therein, a bottom closure to the chamber provided with a threaded opening, said bottom closure carrying film-holding means and at least one channel connecting the opening in said bottom closure with the interior of the chamber.

4. An adapter for reversibly transforming an electron microscope including an electron source and at least one electron lens into an X-ray microscope comprising: in combination, a chamber adapted to be slidably introduced into and accurately positioned in a pole piece of a focusable electron lens of an electron microscope and comprising a clamping means to support an X-ray target References Cited in the file of this patent foil leaving a small aperture alignable with an electron UNITED STATES PATENTS beam passmg longitudinally of the pole piece, with said foil at an electron focus thereby generating X-rays from 2,417,110 11, 1947 a minimum sized source, specimen carrying means to posi- 5 2,650,308 catlln 1953 tion a specimen in the path of the X-rays, and a film holding means to position a photographic film in the OTHER REFERENCES shadow of the specimen, and means to provide for evacu- Cosslett and Nixon: The X-ray shadow microscope, ation of the electron path from the electron source to said Journal of Applied Physics, vol. 24, No. 5, May 1953, foil. 1 10 pp. 616, 623. 

